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Abstract
The global spread and emergence of the plasmid-borne mobile colistin resistance genes (mcr) have threatened the efficacy of colistin, a last-resort antibiotic. It is thought that mcr has a limited prevalence in the USA because colistin has neither been approved nor marketed for use in agriculture. This is important because the rise and dissemination of mcr are widely associated with the misuse of colistin in animal agriculture. However, Hassan and Kassem (2020) predicted that mcr might be introduced to the USA via different routes, including food imports and human travel, since these genes are often carried on highly transmissible plasmids. Here, we hypothesized that mcr and mcr-carrying bacteria are spreading in the USA, despite colistin not being used in agriculture. Additionally, we aimed to identify potential bacterial species that might harbor mcr in the USA and the genomic location of these genes (chromosomal vs. plasmid-borne). Samples were aseptically collected from environmental sources (sewage and surface waters) and imported seafood. Putative colistin-resistant isolates were assessed for their phenotypic resistance profiles using the broth microdilution and Kirby-Bauer disk diffusion assays. Gene-specific PCR analysis was used to screen for the harborage of mcr (mcr-1 to mcr-9). Comprehensive genotypic characterization of the mcr-positive isolates was performed using short-read Illumina® and long-read Oxford Nanopore Technologies (GridION) sequencing. Transformation (heat-shock) and biofilm assays were used to confirm that the mcr genes were plasmid-borne or can persist in the bacterial host, respectively.
mcr-9-positive Morganella morganii (n=1), mcr-9-positive Serratia nevei (n=12), and mcr-3-positive Aeromonas jandaei were detected in the different samples (imported shrimp, influent, effluent and surface water). Notably, mcr-9 was located on transmissible and conjugative IncF and IncHI2 plasmids. Therefore, the findings of this dissertation highlighted the occurrence of mcr in different matrices in the USA for the first time, revealing previously overlooked routes of transmission and identifying unevaluated bacterial hosts that might facilitate the dissemination of these genes. This work calls for the implementation of robust surveillance efforts, especially towards mcr, to help mitigate the spread of the genes across the food supply chain and other resources and preserve the effectiveness of colistin in the USA.